In the identification or quantification of analytes in a sample, often more than one analytical technique is needed to adequately separate the analytes. These techniques generally differ from one another based on the difference in various properties of the analytes. For example, one separation technique may be based on the molecular size of the analytes and another technique may be based on the electrical charge to mass ratios of the analytes. Analytical methods that separate the analytes based on two different analytical techniques are sometimes referred to as "two-dimensional" methods. Analytical techniques suitable for application in separating analytes in liquid samples include, for example, liquid chromatography, electrophoresis, and the like.
Recently, analytical techniques that employ small tubular structures (i.e., microcolumns) have gained wide acceptance. For example, high performance liquid chromatography (HPLC) and capillary electrophoresis (CE) are commonly used techniques for separating analytes, including macromolecules and biomolecules such as proteins, nucleic acids, DNA molecules and fragments, carbohydrates, fatty acids, peptides, and the like. In these systems, a sample that is suspected of containing analytes is sent through a microcolumn. As the molecules in the sample migrate through the microcolumn, depending on the interaction of the analytes with the other substances (such as packing material) in the microcolumn, the analytes separate from one another. Although it is practicable to collect fractions of the liquid exiting a microcolumn and process them through another microcolumn, it is more convenient to couple the microcolumns for fluid transfer. This can obviate the need to collect fractions from one microcolumn and then inject them into another one.
To transfer only portions of a fluid exiting from a microcolumn to another microcolumn, a well designed interface system is important for properly coupling the microcolumns so that the desired liquid portions are transferred without excessive loss. Systems for interfacing a first microcolumn to a second microcolumn have been introduced to enable switching between moving fluid from a first microcolumn and moving fluid from a flush buffer supply to a second microcolumn. For example, U.S. Pat. No. 5,131,991 (Jorgenson et al.) discloses utilizing a valve means connecting the capillary inlet end of a two-dimensional separation system to the chromatography column outlet and to a buffer supply means. The valve means is switchable between a first configuration providing fluid to the capillary inlet end from the buffer supply means and a second configuration providing fluid to the capillary inlet end from the chromatography column. However, in this system, a relatively expensive and sophisticated valve mechanism is needed for switching between the first and the second configurations.
U.S. Pat. No. 5,389,221 (Jorgenson et al.) discloses a combination liquid chromatography and capillary electrophoresis separation system having a flow gating interface. The flow gating interface has an effluent channel and a gating channel (which is shown in FIGS. 1a and 1b). The gating channel 2 transversely intersects the effluent channel 3. Arrows C and D show the flow directions of the gating channel 2 and of the effluent channel 3 respectively. The channel is formed by two plates 4A, 4B stacked together, separated by a gasket (not shown in the drawing) which has a channel cut from it. A liquid chromatography column 5A is connected to the effluent channel 3 upstream portion and an electrophoresis capillary 5B is connected to the effluent channel downstream portion. A flush solution inlet line is connected to the gating channel 2 upstream portion and the flush solution outlet line is connected to the gating channel downstream portion. A valve (not shown in FIGS. 1a and 1b) regulates the flow of flushed solution from the flush solution inlet line to the gating channel upstream portion. The intersection portion is configured so that the rate of flow of the effluent from the effluent channel upstream portion to the effluent channel downstream portion decreases as the rate of flush flow in the gating channel increases. In this system, since plates and gaskets need to be assembled, the manufacturing process is relatively complex. Because precise positioning of the outlet end of the liquid chromatography column and the inlet end of the electrophoresis capillary is important, extensive skill is required to cut the gasket, make the plates, and align the plates with the gasket, the chromatography column, and the capillary. What is needed is a relatively simple and inexpensive flow gating interface. Also needed is a two-dimensional microcolumnar separation apparatus having such a flow gating interface.